调控溶酶体cathepsin B在DNA损伤剂依托泊苷抗前列腺癌中的作用及机制研究

Transformation into hormone-independent prostate cancer and apoptosis resistance to chemotherapy are the major reasons of drug resistance, or metastasis in cancer therapy. Some studies suggest that the content of lysosome cathepsin B in prostate cancer is higher than that of normal tissue. Therefore, the regulation of cathepsin B is expected to be a new way to further improve the efficacy of drugs. The preliminary study found that etoposide induced prostate cancer cells apoptosis in vitro and in vivo, and caused cathepsin B abnormal transport and distribution. Based on this finding, the work will be carried out as follows. Through study on localization, expression and activity of lysosomal cathepsin B induced by etoposide in prostate cancer cells, we reveal the mechanism and signal transduction of cathepsin B activation, and elucidate the material basis of the compound. Based on etoposide activity evaluation in vitro and in vivo, we study the molecular mechanism of regulating cathepsin B expression and activity in prostate cancer cell apoptosis, DNA damage, invasion and metastasis. After etoposide combined with inhibitors, we determine cell survival rate to reveal the relationship between cathepsin B regulation and drug efficacy improvement. Based on the completion of this project, we may provide a new choice in prostate cancer treatment, supply a theoretieal basis for combination drug therapy and offer a new idea to improve chemotherapy of prostate cancer patients.

前列腺癌激素非依赖性转化、抵抗化疗药物诱导的凋亡是导致其耐药、转移的重要原因，有研究提示前列腺癌cathepsin B含量比正常组织高，故调控cathepsin B有望成为提高药物疗效的新途径。申请人前期发现，etoposide体内外诱导凋亡的同时导致前列腺癌cathepsin B异常转运和分布。本项目拟开展：研究etoposide作用于前列腺癌溶酶体cathepsin B的定位、表达和活性，揭示cathepsin B激活机制，阐明化合物作用的物质基础；在体内外活性评价基础上，研究调控cathepsin B表达和活性在etoposide作用于前列腺癌凋亡、DNA损伤、侵袭转移中的机制；通过etoposide联合应用抑制剂揭示调控cathepsin B与提高药物疗效之间的关系，为前列腺癌药物治疗提供新的选择，为联合用药提供理论基础，也为进一步提高前列腺癌化疗疗效提供新的思路。

化疗药etoposide通过诱导DNA损伤和肿瘤细胞凋亡广泛应用于治疗多种癌症。Cathepsin B是溶酶体中的一种组织蛋白酶，在恶性肿瘤细胞的增殖和侵袭转移中起着重要作用。最近研究表明cathepsin B在肿瘤细胞中参与一系列凋亡通路，本课题主要研究cathepsin B在etoposide抗肿瘤中的作用机制。结果显示，在前列腺癌细胞中，etoposide能够引起溶酶体稳定性降低，导致cathepsin B从溶酶体中释放出来，并且活性和表达均增加。为了进一步研究cathepsin B的激活和etoposide诱导的细胞死亡之间的关系，我们采用cathepsin B的抑制剂CA074Me、cathepsin B释放的促进剂，或者应用小干扰RNA敲除cathepsin B的表达、转染过表达载体增加cathepsin B的表达，观察调控cathepsin B对etoposide诱导的细胞死亡的影响，结果表明，etoposide在体内和体外均通过cathepsin B依赖的方式导致DNA损伤和细胞凋亡。另外，cathepsin B参与了etoposide引起DNA损伤的ATM‐Chk2和ATR‐Chk1通路。以上结果表明，cathepsin B在etoposide抗前列腺癌中起着关键作用。该项目的完成，为前列腺癌药物治疗提供新的选择，为联合用药提供理论基础，也为进一步提高前列腺癌化疗疗效提供新的思路。